Mitogen activated protein (MAP) kinases are serine/threonine kinases that mediate signal transduction and orchestrate cellular responses to environmental stress. In mammalian cells, three principle MAP kinase pathways have been identified, including c-Jun N-terminal kinase (JNK), extracellular regulating kinase (ERK), and p38 MAP kinase. p38 is thought to be especially relevant to rheumatoid arthritis (RA) because it is activated in RA synovium and regulates key mediators implicated in the disease. It is regulated by two main upstream MAP kinase kinases, namely MKK3 and MKK6. Despite pre-clinical evidence that p38 blockade should be beneficial in RA, only modest improvement has been noted in clinical trials when patients are treated with p38 inhibitors. This observation might be due to increased activation of ERK and JNK and suppression of anti-inflammatory cytokines like IL-10 by p38 inhibitors. Understanding the mechanisms that limit efficacy is crucial to developing effective new targeted therapies. We now hypothesize that targeting MKK3 or MKK6 might be more effective than traditional p38 inhibitors in diseases like RA. This proposal will explore the biology of MKK3, MKK6, and p38 and test our hypothesis by 1) determining if targeting MKK3 or MKK6 instead of p38 in murine arthritis models preserves IL-10 production and prevents increased activation of other MAP kinases; 2) determining the mechanisms of differential IL-10 regulation and ERK/JNK signaling by MKKs and p38 in cultured macrophages; and 3) determining if p38 blockade suppresses IL-10 and activates ERK/JNK in RA samples and whether this is prevented by targeting MKK3 or MKK6. In addition, a program to evaluate potential MKK3 and MKK6 inhibitors will be initiated to translate the results into a potential therapy. If successful, these studies will support development of novel MAP kinase kinase inhibitors for RA.